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PXD003929

DataSet Summary

  • HostingRepository: PRIDE
  • AnnounceDate: 2017-10-12
  • AnnouncementXML: Submission_2017-10-12_00:06:56.xml
  • DigitalObjectIdentifier:
  • ReviewLevel: Peer-reviewed dataset
  • DatasetOrigin: Original data
  • RepositorySupport: Unsupported dataset by repository
  • PrimarySubmitter: Dennis Goldfarb
  • Title: Identification and Characterization of MCM3 as a KEAP1 Substrate
  • Description: KEAP1 is a substrate adaptor protein for a CUL3-based E3 ubiquitin ligase. Ubiquitylation and degradation of the antioxidant transcription factor NRF2 is considered the primary function of KEAP1; however, few other KEAP1 substrates have been identified. Because KEAP1 is altered in a number of human pathologies and has been proposed as a potential therapeutic target therein, we sought to better understand KEAP1 through systematic identification of its substrates. Towards this goal, we combined parallel affinity capture proteomics and candidate-based approaches. Substrate-trapping proteomics yielded NRF2 and the related transcription factor NRF1 as KEAP1 substrates. Our targeted investigation of KEAP1 interacting proteins revealed MCM3, an essential subunit of the replicative DNA helicase, as a new substrate. We show that MCM3 is ubiquitylated by the KEAP1-CUL3-RBX1 complex in cells and in vitro. Using ubiquitin remnant profiling, we identify the sites of KEAP1-dependent ubiquitylation in MCM3, and these sites are on predicted exposed surfaces of the MCM2-7 complex. Unexpectedly, we determined that KEAP1 does not regulate total MCM3 protein stability or subcellular localization. Our analysis of a KEAP1 targeting motif in MCM3 suggests MCM3 is a point of direct contact between KEAP1 and the MCM hexamer. Moreover, KEAP1 associates with chromatin in a cell cycle-dependent fashion with kinetics similar to the MCM2-7 complex. KEAP1 is thus poised to affect MCM2-7 dynamics or function rather than MCM3 abundance. Together, these data establish new functions for KEAP1 within the nucleus and identify MCM3 as a novel substrate of the KEAP1-CUL3-RBX1 E3 ligase.
  • SpeciesList: scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
  • ModificationList: ubiquitination signature dipeptidyl lysine; phosphorylated residue; acetylated residue; iodoacetamide derivatized residue
  • Instrument: LTQ Orbitrap Velos; LTQ Orbitrap Elite

Dataset History

VersionDatetimeStatusChangeLog Entry
02016-04-08 05:46:29ID requested
12017-10-12 00:06:57announced

Publication List

  1. Mulvaney KM, Matson JP, Siesser PF, Tamir TY, Goldfarb D, Jacobs TM, Cloer EW, Harrison JS, Vaziri C, Cook JG, Major MB, Identification and Characterization of MCM3 as a Kelch-like ECH-associated Protein 1 (KEAP1) Substrate. J Biol Chem, 291(45):23719-23733(2016) [pubmed]

Keyword List

  1. curator keyword: Biological
  2. submitter keyword: HEK293T, SILAC, label free quantification, Di-Gly, APMS

Contact List

    Michael Benjamin Major
    • contact affiliation: Lineberger Comprehensive Cancer Center, Dept of Cell Biology & Physiology, UNC Chapel Hill
    • contact email: ben_major@med.unc.edu
    • lab head:
    Dennis Goldfarb
    • contact affiliation: Cell and Developmental Biology
    • contact email: dennisg@email.unc.edu
    • dataset submitter:

Full Dataset Link List

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  2. PRIDE project URI
Repository Record List
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